The object of this proposal is to study protein-nucleotide interactions in membranes and purified enzymes using recently synthesized, photo-affinity analogs of ZTP and cAMP. The nucleotide photo-affinity analogs to be used in this work have already been utilized by the principal investigator to label ATPases and cAMP binding proteins. Experiments discussed in this proposal will be directed toward the following problems: (1) Optimizing the synthesis and storage of the photo-affinity analogs. (2) Optimizing the use of these reagents as membrane and enzyme active site probes. (3) Labeling the Na, K-ATPase and the divalent metal ion (Mg and Ca) ATPases of human red cell membranes, determining their molecular weights and the number of each enzyme molecule per red cell. (4) Labeling the cAMP binding proteins of human red cell membranes and other membranes. Determination of the number of cAMP binding sites per cell. (5) Labeling the active site of purified Na, K-ATPase and isolating the labeled active site polypeptide. (6) Labeling the regulatory and catalytic sites of cAMP stimulated protein kinase, using the photo-affinity analogs of cAMP and ATP, and isolating the labeled polypeptides. (7) Other ATPase and cAMP requiring systems or enzymes will be tested if time and funds are adequate.